INTRODUCTION

Behavioral processes are paired with patterns of brain activation. Mapping the neural basis of a behavioral distinction, such as visuomotor learning, memory retrieval, associative learning or spatial navigation can be established by controlled multiform experimental manipulation. This can be a change in stimulus information, task instruction or reward and punishment which alter the underlying pattern of brain activation. Monitoring or evoking behavioral data and parallel imaging of neuronal circuits in a living animal’s brain can reveal the correspondence between changes at the cellular and behavioral levels.

SOLUTIONS FOR BEHAVIOR STUDIES

  • Custom-designed microscope configurations with a wide range of accessories
  • Flexible microscope hardware and software connection
  • Synchronization with behavior
  • Input and output signals in a same frame clock
  • Aligned image acquisition and trigger signals

TWO-PHOTON MICROSCOPES FOR BEHAVIOR STUDIES

Femto3D-AcoustoOptic


Femto3D-AcoustoOptic is able to measure neural activity in distributed points of a cubic millimeter scanning volume with an astonishing speed (up to 53 kHz/ROI), which feature makes this microscope unique in 3D two-photon imaging and allowing the measurement of up to 1000 cells simultaneously.




Our Anti-mOtion technology is an acousto-optic scanning method developed for correcting motions appeared during in vivo imaging. This technology extends the points of the random-access point scanning method to fast scanned lines. Based on these lines, scanning is executed on surface or volume elements, containing dendritic segments, spines or cell bodies, with maintained temporal resolution. The scanned parts cover not only the pre-selected regions of interest but also the neighboring areas or volume elements giving an opportunity to decrease the motion artifacts by more than one order of magnitude in behaving animals.

FemtoS


FemtoS was primarily developed to use it for in vivo studies. This system's special feature is the elevated, X-Y-Z moving body which ensures a large space under the objective allowing free and easy positioning of the sample. This feature supports functional behavioral studies of model organisms with different weights and sizes, from mice to even non-human primates while they are moving in a virtual reality environment.

FemtoS-Resonant
With the FemtoS-Resonant you can acquire activity information from the entire field of view with high speed. Recording two-photon fluorescent data from a large field of view makes offline motion correction easy for labelings when the cells can not be identified on the background image without being active.

BEHAVIORAL TOOLS

Virtual reality systems

Virtual Reality (VR) systems enable head-fixed but free-to-move rodents to enter a VR environment and provide a well-controllable experimental protocol to investigate cognition, navigation, learning, memory and operant conditioning, while the animal performs complex behaviors.

Phenosys: JetBall-TFT consists of a TFT surround monitor system, surrounding the animal in 200 degree and a spherical treadmill allowing unobscured field or maze designs. This device can be coupled with optional operant devices, and it allows a restrained animal to navigate in virtual space.

Neurotar: Mobile HomeCage provides a real and familiar moving VR environment. A head-fixed awake rodent walks freely on the flat-floored, air-lifted cage that is moved by the animal’s locomotion, while exploring and navigating during in vivo recordings and imaging experiments.

Femto-Gramophone is an affordable single dimension VR system. The animal runs on a spinning disk while the background on the coupled screen changes according to the speed of the mouse. Gramophone device can be coupled with multiple operant devices such as water reward, airpuff or visual stimulation. The speed of the animal and the two-photon calcium signal can be recorded simultaneously with a built-in module of the main measurement software (MES).

Luigs & Neumann treadmill is a long setup-compatible treadmill, with customized belts up to 4m (13') of length, precise speed and position readout, digital or analog position readout, automatic liquid reward at predefined distance or speed.

Camera for following the behavior


Precisely coupled video recording with two-photon measurement helps to monitor and record the animal's behavior responses given to sensory stimuli simultaneously with neuronal activity. Correlations between functional imaging and behavioral data serves as a highly useful feedback and can extend the result of many in vivo studies.

Head holders

Head holder stands and head plates fix the rodent's head in flexible positions allowing precise measurements in the brain. Head holders are available with different dimensions for mice and rats. For anesthetized rodents, we offer heating pad coupled to the holder, or for behaving animals a stand that ensures the accessibility to jetballs, treadmills or other devices.

Complex bevaior tasks with Bpod module

Bpod is a custom-designed behavior recording system and real-time environment controller for rodent experiments. From high-level programming environments (MATLAB/Python), it provides a low-latency closed-loop link between behavioral events, stimulus delivery and stimulation. Using its built-in liquid reward delivery system, it can be used to power go/no-go discrimination, two alternative force choice and CS/US behavioral paradigms. Bpod data acquisition is fully synchronised with the microscope, and the behavioral events can be precisely aligned to the record of calcium transients.

Digital input-output


MES can drive multiple digital input or output channels to trigger your behavioral devices, record events produced by your behavioral tool, or you can even trigger the start of a measurement with an external trigger input.

IMAGING TOOLS

Green illumination


Green illumination performed by a LED light source allows high-contrast visualization and video camera recording of blood vessels on the surface of many organs by revealing the highly concentrated heme content of the red blood cells which absorbs the light in the spectral region of 300-650 nm. This visualization method helps to navigate in the sample under in vivo conditions.

Intrinsic imaging


The Intrinsic imaging system serves as a monitoring tool before or during in vivo experiments which is optimized to functionally localize brain or spinal cord areas invasively, but without opening the skull. Stimulation evoked signals can be added by Stim Visual software module of MES. Intrinsic imaging is used for many applications, including imaging individual barrels in rodent somatosensory cortex, localizing the visual cortex or the tonotopic organization in auditory cortex.

Software controlled triggering of stimuli


The Stim Visual software module of MES allows loading and sequentially playing videos or images for visual stimulus and synchronizing the stimuli with the evoked neural responses. The stimulation can be triggered and played directly from the measurement computer using MES output signals or from a separate system using any other digital trigger signal. The videos can be played in predefined or in random order. Presenting the stimuli in random order, the software will save the IDs and the timestamps of these videos or pictures, thus the correlation between the stimuli and the acquired calcium data can be studied with ease. In addition, many other stimuli can be triggered such as tactile, whisker, auditory or odor stimuli by software or TTL signals.

Parallel electrophysiology


Electrophysiology module of MES is a convenient tool to acquire and align electrophysiological signals with two-photon imaging data, for both single cell, whole cell patch clamp and multi channel field recording. This module is capable of recording and digitizing your electrophysiological data up to 8 channels. Alternatively electrophysiological data can be imported to MES and synchronized with your imaging data.

Batch processing


Batch processing module of MES software contains efficient tools to analyze entire multi-ROI measurement sets conveniently. It implements the grouped analysis of imaging data at multiple regions and multiple measurement repetitions, and the handling of the resulting calcium transients, electrophysiology time series, and metadata.

DoCallBack


DoCallBack module of MES enables you to call custom MATLAB functions before and after a measurement is executed. Useful for pre-measurement signaling or post-measurement modification of data, annotation, or analysis.

Photostimulation


Behavior can be manipulated via optogenetic methods by inserting light-sensitive receptor proteins into single mammalian neurons in vivo making these neurons sensitive to activation by light of specific wavelengths. Photostimulation of these cells performed by our microscope solutions provides in altering the activity of specific types of neurons to control a subject's behavior.

REFERENCES

Fast 3D Imaging of Spine, Dendritic, and Neuronal Assemblies in Behaving Animals. Gergely Szalay, Linda Judak, Gergely Katona, Katalin Ocsai, Gabor Juhasz, Mate Veress, Zoltan Szadai, Andras Feher, Tamas Tompa, Balazs Chiovini, Pal Maak, Balazs Rozsa, Neuron (2016)


Accurate spike estimation from noisy calcium signals for ultrafast three-dimensional imaging of large neuronal populations in vivo. Thomas Deneux, Attila Kaszas, Gergely Szalay, Gergely Katona, Tamas Lakner, Amiram Grinvald, Balazs Rozsa & Ivo Vanzetta, Nature Communications (2016)


Microglia protect against brain injury and their selective elimination dysregulates neuronal network activity after stroke. Szalay G, Matinecz B, Lenart B, Kornyei Z, Orsolits B, Judak L, Csaszar E, Fekete R, West BL, Katona G, Rozsa B, Denes A, Nature Communications (2016)


Matching Cell Type to Function in Cortical Circuits Luc Estebanez, Jens Kremkow, James F.A. Poulet, Neuron (2015)


In Vivo Monosynaptic Excitatory Transmission between Layer 2 Cortical Pyramidal Neurons. Jean-Sebastien Jouhanneau, Jens Kremkow, Anja L. Dorrn, James F.A. Poulet, Cell Reports (2015)


Synaptic Mechanisms Underlying Sparse Coding of Active Touch Crochet S, Poulet JF, Kremer Y, Petersen CC., Neuron (2011)